Project Summary. Tooth eruption requires alveolar bone resorption and bone formation. We have shown that the osteoclastogenesis needed for resorption of alveolar bone in the coronal region of the bony crypt is regulated by expression of osteoclastogenesis genes in the dental follicle (DF). Similarly, we hypothesize that osteo-inductive genes expressed in the DF, especially those genes upregulated in the basal one-half of the DF, promote tooth eruption by regulating the osteogenesis needed for bone formation that occurs at the base of the alveolar bony crypt. To test this hypothesis, gene microarray studies will be done to determine which osteo-inductive genes are upregulated in the DF at the times of maximal bone growth at the base of the socket. Next, because surgical removal of the basal one-half of the DF prevents tooth eruption and alveolar bone formation, laser capture microdissection coupled with real-time RT-PCR studies will determine which of these osteo-inductive genes are expressed more in the basal portion than in the coronal portion of the DF. Each gene that is upregulated more in the basal portion will then be silenced in vivo using electroporation and RNAi technology specific for the target mRNA of each gene to determine the effect of each gene on eruption times. Regarding osteoclastogenesis, secreted frizzled-related protein-1 (SFRP-1), a molecule that inhibits osteoclastogenesis, is expressed in the DF and it is hypothesized that its gene expression is downregulated by eruption molecules such that osteoclastogenesis can occur. Thus, DF cells will be incubated with molecules that promote osteoclastogenesis for eruption to determine which of them down-regulate SFRP-1 expression. Using osteoclast precursor cells, we also will determine if SFRP-1 inhibits osteoclastogenesis by down- regulating a cell fusion molecule, DC-STAMP. Finally, we hypothesize that stem cells present in the DF might contribute to the osteoclast precursors and osteoblasts needed for eruption. To test this, stem cells isolated from the DF will be incubated with osteoclast-inducing molecules or with osteo-inductive molecules to determine if the stem cells can form osteoclasts or osteoblasts. Relevance. Understanding the molecular regulation of the osteogenesis and osteoclastogenesis needed for eruption may explain why impacted teeth (e.g., 3rd molars) do not erupt, as well as explain the causes of various eruption disorders such as seen in Hutchinson-Gilford Progeria syndrome. To correct eruption disorders, a molecular approach to induce eruption is a long-term goal. Finally, because the periodontal ligament (PDL) is derived from the DF, the molecular regulation of osteoclastogenesis by the DF may suggest a similar role for the PDL in regulating alveolar bone loss seen in periodontitis.